SS18-SSX-mediated Hijacking of mSWI/SNF (BAF) Complexes: Mechanisms and Functional Dependencies Project Summary/Abstract Human synovial sarcoma (SS) is a highly aggressive soft-tissue malignancy which accounts for approximately 10% of all sarcomas. The hallmark molecular lesion which is pathognomonic for the disease is the t(X;18) chromosomal translocation which produces an in-frame fusion between the SS18 gene (Chr18) and one of three genes clustered on the X chromosome, SSX1, SSX2, or SSX4. This event results in a fusion oncoprotein, the SS18-SSX fusion protein, universally considered to be the driver of disease. Several years ago, using biochemical approaches, we discovered SS18 to be a stable, dedicated subunit of the mammalian SWI/SNF (BAF) ATP-dependent chromatin remodeling complex. Further, we discovered that in SS cell lines and model systems, SS18-SSX fusion dominantly incorporates into BAF complexes, displacing wild-type SS18 (the product of the remaining wild-type SS18 allele) as well as the BAF47 (SMARCB1, INI1) subunit likely due to configurational changes and/or steric hindrance. On a functional level, the assembly of the SS18-SSX fusion oncoprotein into BAF complexes results in its de novo targeting on chromatin, particularly to PRC2-repressed loci (such as SOX2), at which BAF complexes oppose PRC2-mediated repression of PRC2 target genes involved in neural development, mesenchymal stem cell maintenance, and others. These biochemistry and functional genomics-centered data provide a powerful foundation for the rigorous interrogation of SS18-SSX-mediated oncogenesis as well as the identification and development of novel therapeutics approaches. In this FusOnC2 Consortium Center for Synovial Sarcoma Biology and Therapeutics, we propose to: (1) Comprehensively define the targets of SS18-SSX-containing BAF complexes genome-wide and determine the relationship of this targeting to SS-specific gene expression, enhancer state, and chromatin topology; (2) Determine the specific feature(s) on histones to which the SSX 78aa tail (and hence the SS18-SSX-bound BAF complex) tethers using biophysical assays; and (3) Discover and mechanistically interrogate synovial sarcoma-specific synthetic lethal vulnerabilities that represent actionable therapeutic strategies. Taken together, each of these three Aims addresses critical questions in the field which currently represent major barriers to both biologic understanding and treatment approaches for this disease. Importantly, successful completion of these aims, in combination with those Aims of Research Projects 2 and 3 within our Center, will inform and enable, in parallel, the development of screening strategies for the identification of novel, synovial sarcoma-specific therapeutics.